PROJECT SUMMARY Glaucoma is a leading source of irreversible blindness worldwide. The disease causes degeneration of retinal ganglion cells (RGCs) and their axons through sensitivity to intraocular pressure (IOP). Many patients continue to lose vision despite efforts to manage IOP. Thus, an unmet clinical need is a treatment that addresses RGC degeneration directly. Our long-term goal is to address this need by identifying new therapeutic targets based on neuronal repair, protection and restoration. Consistent with this goal, in the current cycle we discovered a powerful form of intrinsic RGC protection involving the TRPV1 (transient receptor potential vanilloid-1) cation channel. We found that RGC spontaneous excitation and axon signaling is enhanced in response to elevated IOP through up-regulation, translocation and increased activation of TRPV1. Silencing TRPV1 by knock-out (Trpv1-/-) or pharmacological antagonism eradicates this enhancement, raises the threshold for RGC axon signaling, and accelerates by two-fold axon degeneration in our inducible model. Our objective now is to investigate how TRPV1 exerts this compensatory influence in RGCs and whether TRPV1-mediated enhancement could be harnessed therapeutically in glaucoma by increasing RGC resistance to stress. To do so, we will test the central hypothesis that in glaucoma, increased TRPV1 in RGC dendrites stabilizes cytoskeletal and synaptic structures by potentiating glutamatergic signaling, resulting in enhanced excitation. This hypothesis is supported by published results from the current cycle that form the premise for this continuation. While TRPV1 expression is normally low, we found that elevated IOP induces a transient increase in RGC TRPV1 mRNA and protein. This up-regulation involves translocation of TRPV1 to RGC dendrites proximal to glutamatergic synapses, where it supports focal increases in Ca2+ and amplified depolarizing currents. These changes parallel the IOP-induced enhancement of RGC spontaneous axon activity that is absent in Trpv1-/- mice. To link these observations mechanistically and test their therapeutic value, we will combine physiological, cell-imaging and molecular tools to probe TRPV1 function in RGCs using inducible (microbead occlusion) and chronic (DBA2J mouse) models of glaucoma. Experiments in Aim 1 will test how TRPV1 influences RGC dendrite and synapse survival during progression and whether this influence differs between RGC types identified physiologically and morphologically. Aim 2 will measure TRPV1's influence on glutamatergic signaling in RGC types, its interactions with synaptic and dendritic proteins, and whether its actions in glaucoma depend upon activation of the mechanosensitive TRPV4 subunit. Finally, Aim 3 will probe whether manipulating TRPV1 activation and expression boosts excitatory enhancement, prevents degeneration, and improves visual performance. These innovative studies offer a new understanding of progression in glaucoma at an unprecedented level of detail with the goal for new therapies based on manipulating neuronal physiology and signaling.